| Cadmium (Cd) is accumulated in soils mainly due to anthropogenic activities such as mining,urban traffic and fertilizer application. Cd is a nonessential element and posses highly toxic to plants. It can be absorbed by plants and eventually enters the human body through the food chain, thereby Cd constitutes a serious threat to human health. Phytoremediation is an efficient biotechnology to absorb Cd from the soil and to accumulate it in aerial parts of plants which can be easily harvested. The usage of poplar species has been proposed for candidates of phytoremediation because they are characterized by deep-rooting systems, fast-growth and large-biomass. However, different clones of the same poplar species may exhibit variable Cd tolerance due to intraspecificgenetic dissimilarity. Besides,a holistic view on physiological and molecular mechanisms in response to Cd exposure has so far not been reported. Here, we used poplar species as materials to explore the variation in Cd accumulation, translocation and tolerance among seven poplars; subesequently, the physiological and molecular mechanism of Cd uptake, translocation and accumulation were fully studied. Finally, the mechanism of Cd uptake and tolerance in poplar over-expressed y-glutamylcysteine synthetase was explored.The main results were as follow:1. Cadmium tolerance among different poplar speciesSelection of poplar species with greater Cd tolerance and exploiting the physiological mechanisms involved in Cd tolerance are crucial for application of these species to phyto-remediation. The aim of this study is to investigate variation in Cd tolerance among the seven poplar species. Cuttings of seven poplar species were cultivated for10weeks before exposure to either0or200μM CdSO4for20days.Gas exchange in mature leaves was determined by a portable photosynthesis system. Cd concentrations in tissues were analyzed by a flame atomic absorbance spectrometry. Subsequently, Cd amount per plant, bio-concentration factor (BCF) and translocation factor (Tf) were calculated. Cd exposure caused decline in photosynthesis in five poplar species. Among the seven species, P.×canescens displayed the highest Cd concentrations, the largest Cd amount and the highest BCF in aerial parts, and Tf under Cd exposure. Under Cd stress, increases in total soluble sugars in bark and leaves, and increases in starch in roots and leaves of P.×canescens were found. Based on results of this pot experiment, it is concluded that P.×canescens is more tolerance to Cd and superior to other six species for Cd phyto-remediation.2.Net cadmium flux and the physiological responses to different Cd exposure time in Populus×canescensTo characterize the dynamics of Cd2+flux in the rhizosphere and to study Cd plant-internal partitioning in roots, wood, bark and leaves in relation to energy metabolism, reactive oxygen species (ROS) formation and antioxidants, Populus×canescens plantlets were exposed to0or50μM CdSO4for up to20days. A strong net Cd2+influx in root apex was observed after Cd exposure for24h, even if net Cd+influx decreased gradually in roots. A large amount of Cd was accumulated in roots. Cd ions were uploaded via the xylem to leaves and further transported to the phloem where significant accumulation was detected. Cd accumulation led to decreased photosynthetic carbon assimilation but not to the depletion in soluble carbohydrates. Increased levels of ROS were present in all tissues, except for the bark of Cd exposed poplars. To combat Cd induced O2·-and H2O2, P. x canescens appeared to rely mainly on the formation of soluble phenolics as these compounds showed the highest accumulation in the bark and the lowest in wood. Other potential radical scavengers such as proline, sugar alcohols and antioxidant enzymes showed tissue-and exposure time-specific responses to Cd.3. A transcriptomic network underlies microstructural and physiological responses to cadmium in Populus×canescensBark tissue of Populus×canescens can hyperaccumulate Cd, but microstructural, transcriptomic and physiological response mechanisms are poorly understood. Histochemical assays, transmission electron microscopic observations, energy-dispersive X-ray microanalysis, transcriptomic and physiological analyses have been performed to enhance our understanding of Cd accumulation and detoxificationin P.×canescens. Cd was allocated to the phloem of the bark and subcellular Cd compartmentalization occurred mainly in vacuoles of phloem cells. Transcripts involved in microstructural alteration, changes in nutrition and primary metabolism, and stimulation of stress responses showed significantly differential expression in the bark of P.×canescens exposed to Cd. About48%of the differentially regulated transcripts formed a co-regulation network in which43hub genes played a central role both in cross talk among distinct biological processes and in coordinating the transcriptomic regulation in the bark of P.×canescens in response to Cd. The Cd transcriptome in the bark was mirrored by physiological readouts. Cd accumulation led to decreased total N, P, Ca and increased S in the bark. Cd inhibited photosynthesis, resulting in decreased carbohydrate levels. Cd induced oxidative stress and antioxidants including free proline, soluble phenolics, ascorbate and thiol compounds. These results suggest that an orchestrated microstructural, transcriptomic, and physiological regulation may sustain Cd hyperaccumulation in P.×canescens bark and provide new insights into engineering woody plants for phytoremediation.4. Over-expression of y-glutamylcysteine synthetase mediates changes in cadmium influx, allocation, and detoxification in poplarOverexpression of y-glutamylcysteine synthetase in the cytosol of Populus tremula x P. alba displayed higher total thiols and glutathione concentrations in leaves than wildtype poplars, holding a potential for remediating cadmium (Cd) contaminated soil. However, net Cd2+influx, Cd tolerance, and the underlying molecular and physiological mechanisms remain to be elucidated in these plants. Plants of wildtype and transgenic poplars grown in hydroponics were exposed to0or100μM Cd for80days. The poplar plants displayed the highest net Cd2+influxes into roots at pH5.5and0.1mM Ca2+, with higher Cd2+uptake rates in the transgenics than in the wildtype. Consistently, induced transcript levels of several genes involved in Cd transport and detoxification, such as PCS, ZIP6.2, NRAMP1.3and PCR2, were detected in roots of transgenic compared to wildtype plants under control and Cd exposure conditions. Transgenics displayed more Cd accumulation in aerial parts, less decrease in net photosynthesis (4) and biomass accumulation, higher concentrations of soluble sugars and starch than wildtype plants in response to Cd exposure. Moreover, transgenic plants showed lower concentrations of O·-and H2O2, higher concentrations of total thiols, GSH and GSSG in roots and leaves, elevated levels of soluble phenolics and free proline, and stimulated foliar GR activity compared to wildtype plants. These results indicate that transgenics are more tolerant to Cd than wildtype plants, which is probably resulted from GSH-mediated induction of transcript levels of genes involved in Cd transport and detoxification. |
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